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Examination of Thymic Positive and Negative Selection by Flow Cytometry
Published on: October 8, 2012
Ly-49 antigen defines an alpha beta TCR population in i-IEL with an extrathymic maturation
1Département d'Immunologie Institut Pasteur (UA CNRS 359), Paris, France.
In this paper we describe a monoclonal antibody, JR9-318, which was obtained from a wild-derived mouse immunized with the M14T cell line (CD3+CD4-CD8-) in order to describe antigenic molecules expressed during T cell differentiation. This mAb precipitated a protein from the M14T cell line or subclones that migrates like a T cell receptor heterodimer at 85-90 kDa under non-reducing conditions and 45-50 kDa under reducing conditions. The molecular characteristics of this antigen and the data obtained by sequential immunoprecipitation led us to conclude that JR9-318 mAb recognizes the Ly-49 molecule already described. Using direct immunofluorescence staining, this mAb was shown to identify a small population of cells in the adult thymus but not in the spleen, lymph node or bone marrow of most laboratory mouse strains. The phenotypic characteristics of this thymic subpopulation do not correspond to those of any other known classical population. A more prominent population of CD3+ alpha beta+ T cells is stained by this mAb in intestinal intra-epithelial lymphocytes with a peculiar phenotype, different from the one found in the thymus. These cells express V beta families usually deleted in the thymus of BALB/c mice. We discuss here the origin of these T cell subpopulations and the possible maturation of T cells without thymic influence.
In this paper we describe a monoclonal antibody, JR9-318, which was obtained from a wild-derived mouse immunized with the M14T cell line (CD3+CD4-CD8-) in order to describe antigenic molecules expressed during T cell differentiation. This mAb precipitated a protein from the M14T cell line or subclones that migrates like a T cell receptor heterodimer at 85-90 kDa under non-reducing conditions and 45-50 kDa under reducing conditions. The molecular characteristics of this antigen and the data obtained by sequential immunoprecipitation led us to conclude that JR9-318 mAb recognizes the Ly-49 molecule already described. Using direct immunofluorescence staining, this mAb was shown to identify a small population of cells in the adult thymus but not in the spleen, lymph node or bone marrow of most laboratory mouse strains. The phenotypic characteristics of this thymic subpopulation do not correspond to those of any other known classical population. A more prominent population of CD3+ alpha beta+ T cells is stained by this mAb in intestinal intra-epithelial lymphocytes with a peculiar phenotype, different from the one found in the thymus. These cells express V beta families usually deleted in the thymus of BALB/c mice. We discuss here the origin of these T cell subpopulations and the possible maturation of T cells without thymic influence.
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